Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:3.6.1.3 (ATPase)
65,361 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Spin-label EPR spectroscopy of shark rectal gland Na,K-ATPase modified at cysteine residues with a variety of maleimide-nitroxide derivatives is used to characterize the different classes of sulphydryl groups. The spin-labelled derivatives vary with respect to charge and lipophilicity, and the chemical reactivity towards modification and inactivation of the Na,K-ATPase is dependent on these properties. Ascorbate is used to reduce the spin-labels in situ, and the kinetics of reduction of the protein-bound spin-labels are found also to depend on the nature of the maleimide-nitroxide derivative. The Na,K-ATPase is labelled either at Class I groups (with retention of enzymatic activity) or at Class II groups (where the enzymatic activity is lost). Although Class I groups are labelled more readily than are Class II groups they are only slightly more susceptible to reduction by ascorbate than the Class II groups, indicating no major difference in environment. The spectral difference observed between immobilized and mobile spin-labels with both Class I and Class II groups labelling is not reflected in widely different reduction kinetics for these two spectral components. Solubilization of the enzyme in an active form does not change the protein structure in terms of increased accessibility of the SH-groups to reduction by ascorbate. The results are discussed in terms of the location of the different SH-groups and the origins of the differences in mobility evident in the EPR spectra of the spin-labelled SH-groups.
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PMID:Analysis of thiol-topography in Na,K-ATPase using labelling with different maleimide nitroxide derivatives. 133 3

Isolated kidney cells accumulated L[1-14C]ascorbic acid in a time-dependent manner and reached a steady state after 15 min at 37 degrees C. Initial velocity for uptake was over 300 pmol/mg protein per min when cells were separated from the bathing solution using a density gradient established during centrifugation. The uptake process was saturable with an apparent concentration at half maximal uptake of 36 mumols/L. Ascorbate uptake was reduced by metabolic inhibitors and was temperature dependent. Although ascorbic acid is an acid anion at pH 7.4, uptake did not appear to be inhibited by other acid anions such as p-aminohippurate and probenecid; however, involvement of the ion gradient established by Na+, H(+)-adenosine triphosphatase could not be confirmed. Replacing the sodium ion with other monovalent ions reduced the accumulation of ascorbate significantly. Isoascorbic and dehydroascorbic acids inhibited ascorbate uptake (34 and 13 mmol/L, respectively), whereas high concentrations of glucose showed some stimulation. These findings indicated that ascorbic acid is reabsorbed by the kidney in a sodium-dependent active transport process that is not common to other acid anions and has some specificity for the ascorbic acid structure.
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PMID:Characterization of ascorbic acid uptake by isolated rat kidney cells. 199 58

When 5-methylphenazinium methylsulfate and a reductant (ascorbate or NADH) are added together to a suspension of resealed chromaffin-vesicle membranes, the pH gradient (inside acidic) and the membrane potential (inside positive) established by the H(+)-translocating adenosine triphosphatase (ATPase) are rapidly dissipated. Dissipation of the pH gradient may be observed using either the optical probe acridine orange or the weak base methylamine. Dissipation of the membrane potential may be observed using the potential-dependent dye oxonol VI. A reductant and 5-methylphenazinium methylsulfate added in combination will also abolish a K+ diffusion potential across chromaffin-vesicle membranes but not across liposome membranes. 5-Methylphenazinium methylsulfate oxidizes cytochrome b561 in chromaffin-vesicle ghosts. Ascorbate readily reduces cytochrome b561, but reduction of cytochrome b561 by NADH is greatly enhanced in the presence of 5-methylphenazinium methylsulfate. These results are consistent with a mechanism in which proton gradient dissipation (a net efflux of H+) is caused by an influx of electrons through the membrane-protein cytochrome b561 coupled with an efflux of H carried by the reduced species 5-methyl-10-hydrophenazine. Although 5-methylphenazinium has been thought to accumulate within acidic vesicles as a weak base, this accounts for neither proton gradient dissipation nor for intravesicular accumulation of the compound.
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PMID:5-Methylphenazinium methylsulfate mediates cyclic electron flow and proton gradient dissipation in chromaffin-vesicle membranes. 221 89

A 3-fold higher concentration of "endogenous digitalis" is found in the serum of patients with essential hypertension than in the serum of normotensives, whose concentration was determined in 22 normotensive probands by an receptor assay using isolated (Na+ + K+)-ATPase as 76.3 +/- 9.3 nM ouabain equivalents. Since the concentration of "endogenous digitalis" was 7-19 fold higher in 2 patients, who had become uremic due to a malignant hypertension and since their serum levels fell 3-fold by hemodialysis, the hemofiltrate was used for partial purification of the substance. This was possible by hydrophobic chromatography on Amberlite XAD-2, octadecyl-coated silica gel, anion exchange chromatography and affinity chromatography on membrane-bound (Na+ + K+)-ATPase. The partially purified substance behaved like the material described by Cloix et al. (1985) Biochem. Biophys. Res. Commun. 131:1234-1240 and competed with digoxin for digoxin antibodies. Ascorbic acid isolated on the search for an inhibitor of (Na+ + K+)-ATPase from beef brain inhibited [3H]ouabain binding due to a decrease of the ouabain binding sites by a reduction of a group within the ATP binding site of the enzyme. Unsaturated fatty acids claimed to be "endogenous digitalis (Tamura et al. [1985] J. Biol. Chem. 260:9672-9677)" also lowered the capacity of the cardiac glycoside binding site but did not compete with ouabain.
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PMID:Sodium pump inhibitor in the serum of patients with essential hypertension and its partial purification from hemofiltrate. 243 45

1. Ascorbic acid, diamide and N-ethylmaleimide inhibit Na+ + K+-ATPase activity in toad corneal epithelium. 2. Ascorbic acid, diamide and N-ethylmaleimide increase alpha-aminoisobutyric acid accumulation in this tissue. 3. The effects of these compounds on corneal amino acid and ion transport are not mediated through alterations in Na+ + K+-ATPase activity.
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PMID:Na+ + K+-ATPase activity and transport processes in toad corneal epithelium. 288 79

Ascorbic acid and Mg-ATP were found to regulate norepinephrine biosynthesis in intact secretory vesicles synergistically and specifically, using the model system of isolated bovine chromaffin granules. Dopamine uptake into chromaffin granules was shown to be unrelated to the presence of Mg-ATP and ascorbic acid at external dopamine concentrations of 7.5 and 10 mM. Under these conditions of dopamine uptake, norepinephrine biosynthesis was enhanced 5-6-fold by Mg-ATP and ascorbic acid compared to control experiments with dopamine only. Furthermore, norepinephrine formation was enhanced approximately 3-fold by ascorbic acid and Mg-ATP together compared to norepinephrine formation in granules incubated with either substance alone. The action of Mg-ATP and ascorbic acid together was synergistic and independent of dopamine content of chromaffin granules as well as of dopamine uptake. The apparent Km of norepinephrine formation for external ascorbic acid was 376 microM and for external Mg-ATP was 132 microM, consistent with the larger amounts of cytosolic ascorbic acid and ATP that are available to chromaffin granules. Other physiologic reducing agents were not able to increase norepinephrine biosynthesis in the presence or absence of Mg-ATP. In addition, maximum enhancement of norepinephrine biosynthesis occurred only with the nucleotide ATP and the cation magnesium. The mechanism of the effect of ascorbic acid and Mg-ATP on norepinephrine biosynthesis was investigated and appeared to be independent of a positive membrane potential. The effect was also not mediated by direct action of ADP, ATP, or magnesium on the activity of soluble or particulate dopamine beta-monooxygenase. These data indicate that Mg-ATP and ascorbic acid specifically and synergistically co-regulate dopamine beta-monooxygenase activity in intact chromaffin granules, independent of substrate uptake. Although the mechanism is not known, the data are consistent with the possibility that the chromaffin granule ATPase mediates these effects.
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PMID:Ascorbic acid and Mg-ATP co-regulate dopamine beta-monooxygenase activity in intact chromaffin granules. 314 26

The ATPase (EC 3.6.1.3) of sarcoplasmic reticulum vesicles was reacted to various extents with thiol-directed spin labels. By suspension of the preparation in appropriate solutions, the enzyme could be placed and held in certain intermediate states of the ATPase cycle, or it could be set into steady-state catalysis. Ascorbate added to the system destroyed the spin-label signals with undetectable distortion of the electron paramagnetic resonance spectrum. In general, in the presence of ascorbate, undestroyed signal as a function of time could be described as the sum of two first-order reductions going on in separate compartments with different ascorbate concentrations. In different enzymatic states the proportion of total signal in the two compartments was different, but the first-order velocity constants remained the same. If the labeled membrane was first attacked with Triton, then exposed to ascorbate, signal was destroyed according to a single first-order constant, equal to the faster of the two constants observed with intact membrane, and equal to the constant whereby ascorbate attacks free label in solution. The data were reconciled by a simple rotary model, envisioning that an enzymatic state corresponds to an average angular position of the ATPase and thereby determines the proportion of labeled thiols exposed to external and internal ascorbate concentrations.
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PMID:Change in state of spin labels bound to sarcoplasmic reticulum with change in enzymic state, as deduced from ascorbate-quenching studies. 437 32

Cultures of osteoblastlike cells obtained from the endosteal surfaces of rabbit long bones formed and mineralized an extracellular matrix when they were supplied daily with medium containing fresh ascorbate. No matrix formed without this supplementation. The matrix mineralized whether or not beta-glycerophosphate, a substrate of alkaline phosphatase, was added to the medium. The ion-transporting ATPase activities of untreated, ascorbate-treated, and ascorbate plus beta-glycerophosphate-treated cells were measured. Ascorbate-treated and ascorbate plus beta-glycerophosphate-treated cells had similar enzyme activities. The activities of the Ca2+-ATPase; Ca2+,Mg2+-ATPase; and alkaline phosphatase in treated cells were elevated over the activities in untreated cells. Na+,K+-ATPase activity was lower in treated than in untreated cells. HCO3--ATPase activity was not changed by treatment. Alkaline phosphatase activity was 20 times higher in freshly isolated osteoblastlike cells than in cells grown to confluence in primary culture. In addition, subculturing further reduced the activity of this osteoblast-marker enzyme. The activities of the ion-transporting ATPases and alkaline phosphatase in second passage cells were similar to the activities of these enzymes in fresh, noncalcifying tissues. Nevertheless, second passage cells retain the ability to mineralize an extracellular matrix, and their ion-transporting ATPase and alkaline phosphatase activities are altered when the cells mineralize a matrix.
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PMID:Ion-transporting ATPases and matrix mineralization in cultured osteoblastlike cells. 609 28

Vanadyl (VO2+) is a potent inductor of the lipid peroxidation in brain microsomes. This effect, however, is obtained at concentrations by two orders of magnitude higher (10(-4)-10(-3)M) than those which effectively inhibit the brain microsomal Na,K-ATPase. At 10(-6)M VO2+ which inhibits 50% of the Na,K-ATPase activity there is no measurable malonyldialdehyde production. Vanadate (VO-3) which is an equally potent inhibitor of Na,K-ATPase as VO2+ has almost no capacity to induce the lipoperoxidation. The addition of 10(-4)M ascorbate to the brain microsomes stimulates the lipoperoxidation to the maximum level regardless of the presence or absence of exogenous vanadium ions. Ascorbate-induced inhibition of brain Na,K-ATPase which is known to be associated with lipoperoxidation is strictly additive with the vanadyl (VO2+) inhibition of this enzyme. Even at submaximal concentrations there is no indication for any potentiation between these two inhibitory systems. The disparity between the mechanisms of ascorbate and vanadyl-induced inhibition of Na,K-ATPase is also documented by the effect of EDTA which inhibits the former type only. It is concluded, that the vanadium-induced inhibition of brain microsomal Na,K-ATPase is not related to induction of lipoperoxidative capacity of the brain.
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PMID:Vanadyl (VO2+) induced lipoperoxidation in the brain microsomal fraction is not related to VO2+ inhibition of Na,K-ATPase. 614 42

Using EDTA and illuminated bovine retinas homogenate it has been found that Na+-K+-ATPase was stimulated by noradrenaline (NA), dopamine (DA), cAMP and dibutyryl cAMP. Phentolamine, an alpha adrenergic antagonist and INPEA, a beta adrenergic antagonist used at the same concentrations as catecholamines (CA) depressed the stimulatory effect of NA. Phentolamine increased the stimulatory effect of DA but INPEA inhibited it. It is proposed, therefore, that DA acts on retina Na+-K+-ATPase by putative DA receptors regulated by adrenergic antagonists. Ascorbic acid (AA), 0.125-1.0 mM depressed Na+-K+-ATPase activity of retinas, rat brain cortex and cerebellum homogenates. The results are consistent with the hypothesis that CA and AA can exert their effects in the presence of EDTA. In the retina the effects of CA is mediated by cAMP and alpha receptor influencing process. AA is postulated as a factor regulating the stimulation of CA on Na+-K+-ATP-ase and thereby the turnover of neurotransmitters in the central nervous system tissues.
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PMID:Possible role of ascorbic acid in catecholamines-stimulated Na+-K+-adenosine triphosphatase activity of the central nervous tissues. 615 50


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